基于生物芯片检测结直肠癌相关基因的甲基化

Screening for Epigenetically Masked Genes in Colorectal Cancer by Microarray

【目的】应用微矩阵基因芯片技术筛选结直肠癌甲基化基因。【方法】分别提取6对结直肠癌组织和癌旁正常组织DNA,超声破碎及甲基化DNA富集后与甲基化芯片(Nimble Gen human DNA Methylation 3×720K Cp G)进行杂交,杂交信号经扫描等处理后采用生物信息学对检测结果进行分析。并对筛选出来的基因进行验证。【结果】一共发现有2 296个高甲基化基因,其中有293个基因无文献报道。生物信息学分析显示,所有的甲基化基因随机分布在24对染色体上。聚类分析结果显示,某些在细胞分裂和肿瘤发生发展中起重要作用的生理过程中存在大量甲基化差异基因,如DNA修复,细胞周期和侵袭等。初步验证发现,Opmcl基因在结直肠癌组织及癌旁正常组织中存在甲基化差异。【结论】在结直肠癌中运用DNA甲基化芯片进行甲基化基因筛查是一种有效的方法。

[Objective] We used the DNA methylation microarrays to investigate the hypermethylated genes in colorectal cancer and adjacent normal mucosa. [Method] DNA was extracted from six pairs of CRC and adjacent normal mucosa and sonicated into fragments ranging in size from 300-1000 bp, followed by methylated DNA Immunoprecipitation analysis and then sent to NimbleGen for Microarray hybridization according to their standard protocol. The hybridized array is then scanned and the resulting image analyzed. The biologic functions were analyzed by bioinformatics methods. In addition, preliminary validation studies were done in six pairs of samples by MSP （methylation-specific PCR）. [ Results] Compared with the normal tissues, there are 2 296 genes were hypermethylated, in which .293 hypermethylated genes were unreported. All these genes were randomly distributed on all the chromosomes. According to gene ontology analysis, the genes involved in some physiological processes which play important roles in the cell division and the development of tumor are methylated, such as DNA repair, cell cycle and invasion etc. Preliminary validation, the Opcml in thirty top-ranking genes was shown hypermethylated in six pairs of colorectal cancer （CRC） and adjacent normal mucosa. [ Conclusions ] High density DNA methylation microarrays is an effective method for screening aberrantly methylated genes in CRC.